T-lymphocyte migratory circuits in human and nonhuman primates remain largely unexplored due to the difficulty of defining cell trafficking in vivo. However, this knowledge may reveal critical aspects of immunity and T-lymphocyte homeostasis in both health and disease. Furthermore, in vivo T-lymphocyte trafficking studies may facilitate defining mechanism(s) of immune dysfunction in the nonhuman primate model for acquired immunodeficiency syndrome (AIDS). Here, we developed a model for in vivo T-lymphocyte trafficking in nonhuman primates, and delineated homing characteristics of unstimulated peripheral blood mononuclear cells (PBMCs) to lymphoid and nonlymphoid compartments in healthy rhesus macaques. T-lymphocyte homing of autologous, carboxyfluorescein diacetate succinimidyl ester (CFSE)-labeled PBMCs was defined within 48 h of intravenous transfer. The highest relative frequency of CFSE+ T lymphocytes was observed in peripheral blood and spleen. Expression of chemokine receptor CCR7 and its ligands correlated with recirculation of T lymphocytes through the periphery and homing to paracortical regions of lymph node, where cells remained largely excluded from B-cell follicles. T-lymphocyte trafficking was also detected to the liver and bone marrow, and at low levels to the thymus and small intestine. The liver contained the highest proportion of CD45RA- T lymphocytes, consistent with homing of activated/memory T lymphocytes to this nonlymphoid site. Our data suggest that lymphoid and nonlymphoid organs are under continuous immunosurveillance in healthy macaques, and that this model may serve to investigate aberrant patterns in disease.